Combined Gap-Polymerase Chain Reaction and Targeted Next-Generation Sequencing Improve α- and ß-Thalassemia Carrier Screening in Pregnant Women in Vietnam.

Vietnam has a high thalassemia burden. We collected blood samples from 5880 pregnant Vietnamese women during prenatal health checks to assess thalassemia carrier frequency using combined gap-polymerase chain reaction (gap-PCR) and targeted next-generation sequencing (NGS). Thalassemia carriers were identified with prevalence of 13.13% (772), including 7.82% (460) carriers of α-thalassemia (α-thal), 5.31% (312) carriers of ß-thalassemia (ß-thal), and 0.63% (37) concurrent α-/ß-thal carriers. Deletional mutations (368) accounted for 80.0% of α-thal carriers, of which, --SEA (Southeast Asian) (n = 254; 55.0%) was most prevalent, followed by the -α3.7 (rightward) (n = 66; 14.0%) and -α4.2 (leftward) (n = 45; 9.8%) deletions. Hb Westmead (HBA2: c.369C>G) (n = 53) and Hb Constant Spring (Hb CS or HBA2: c.427T>C) (in 28) are the two most common nondeletional α-globin variants, accounting for 11.5 and 6.0% of α-thal carriers. We detected 11 different ß-thal genotypes. Hb E (HBB: c.79G>A) (in 211) accounted for 67.6% of ß-thal carriers. The most common ß-thal genotypes were associated with mutations at codon 17 (A>T) (HBB: c.52A>T), codons 41/42 (-TTCT) (HBB: c.126_129delCTTT), and codon 71/72 (+A) (HBB: c.217_218insA) (prevalence 0.70%, 0.68%, and 0.2%, respectively). Based on mutation frequencies calculated in this study, estimates of 5021 babies in Vietnam are affected with clinically severe thalassemia annually. Our data suggest a higher thalassemia carrier frequency in Vietnam than previously reported. We established that combining NGS with gap-PCR creates an effective large-scale thalassemia screening method that can detect a broad range of mutations.

Detection of maternal carriers of common α-thalassemia deletions from cell-free DNA.

α-Thalassemia is a common inherited blood disorder manifested mainly by the deletions of α-globin genes. In geographical areas with high carrier frequencies, screening of α-thalassemia carrier state is therefore of vital importance. This study presents a novel method for identifying female carriers of common α-thalassemia deletions using samples routinely taken for non-invasive prenatal tests for screening of fetal chromosomal aneuploidies. A total of 68,885 Vietnamese pregnant women were recruited and α-thalassemia statuses were determined by gap-PCR, revealing 5344 women (7.76%) carried deletions including αα/--SEA (4.066%), αα/-α3.7 (2.934%), αα/-α4.2 (0.656%), and rare genotypes (0.102%). A two-stage model was built to predict these α-thalassemia deletions from targeted sequencing of the HBA gene cluster on maternal cfDNA. Our method achieved F1-scores of 97.14-99.55% for detecting the three common genotypes and 94.74% for detecting rare genotypes (-α3.7/-α4.2, αα/--THAI, -α3.7/--SEA, -α4.2/--SEA). Additionally, the positive predictive values were 100.00% for αα/αα, 99.29% for αα/--SEA, 94.87% for αα/-α3.7, and 96.51% for αα/-α4.2; and the negative predictive values were 97.63%, 99.99%, 99.99%, and 100.00%, respectively. As NIPT is increasingly adopted for pregnant women, utilizing cfDNA from NIPT to detect maternal carriers of common α-thalassemia deletions will be cost-effective and expand the benefits of NIPT.

The contribution of form repetition to listeners' expectation of givenness in online reference resolution.

Although it is clear that unaccented referring expressions are associated with given information in a discourse (Dahan et al., 2002), it is less clear what aspects of givenness are relevant. We examine whether listeners' expectation of givenness depends on repetition of a referring expression or on contextual evocation of a referent. The results from two visual world eye-tracking experiments suggest that for interpretation, listeners associated reduced prominence with a repeated referring expression. Listeners expect previously evoked referents to be candidates for reduced referring expressions only when they are referred to with the exact same referential form. The data also suggest that when referents are referred to with different referential forms across utterances, accenting facilitates linking those forms for co-reference.

Assessing the virulence of Cryptococcus neoformans causing meningitis in HIV infected and uninfected patients in Vietnam.

We previously observed a substantial burden of cryptococcal meningitis in Vietnam atypically arising in individuals who are uninfected with human immunodeficiency virus (HIV). This disease was associated with a single genotype of Cryptococcus neoformans (sequence type [ST]5), which was significantly less common in HIV-infected individuals. Aiming to compare the phenotypic characteristics of ST5 and non-ST5 C. neoformans, we selected 30 representative Vietnamese isolates and compared their in vitro pathogenic potential and in vivo virulence. ST5 and non-ST5 organisms exhibited comparable characteristics with respect to in vitro virulence markers including melanin production, replication at 37°C, and growth in cerebrospinal fluid. However, the ST5 isolates had significantly increased variability in cellular and capsular sizing compared with non-ST5 organisms (P < .001). Counterintuitively, mice infected with ST5 isolates had significantly longer survival with lower fungal burdens at day 7 than non-ST5 isolates. Notably, ST5 isolates induced significantly greater initial inflammatory responses than non-ST5 strains, measured by TNF-α concentrations (P < .001). Despite being generally less virulent in the mouse model, we hypothesize that the significant within strain variation seen in ST5 isolates in the tested phenotypes may represent an evolutionary advantage enabling adaptation to novel niches including apparently immunocompetent human hosts.

Compliance to dietary guidelines on fruit and vegetable intake and prevalence of hypertension among Vietnamese adults, 2015.

AIMS: The aim of this study was to investigate whether the Western dietary guidelines on fruit and vegetable intake are associated with blood pressure parameters and hypertension among Vietnamese adults. METHODS: Participants included 1384 women and 1049 men aged 18-69 years from the 2015 Vietnam national survey on risk factors of non-communicable diseases. Associations between dietary intake score based on the Dietary Approaches to Stop Hypertension (DASH) guidelines and World Health Organization recommendations on fruit and vegetable consumption and blood pressure parameters and hypertension were evaluated by multivariate regression analyses. RESULTS: Approximately 17.0% and 40.1% of participants met the respective definitions of hypertension according to Joint National Committee 7 (JNC7) and 2017 American College of Cardiology/American Heart Association (ACC/AHA) Hypertension Guideline. Highest tertiles of DASH scores for fruit intake were significantly associated with increased blood pressure parameters, particularly in women. Hypertension was associated with DASH score for fruit intake with odds ratios and 95% confidence intervals for tertiles 2-3 versus tertile 1: 1.31 (0.98, 1.76) and 1.43 (1.05, 1.93) for JNC7; 1.26 (1.01, 1.58) and 1.31 (1.04, 1.66) for 2017 ACC/AHA guideline (all p-trend <0.05). No association with blood pressure parameters and hypertension was observed for DASH score for vegetable intake and meeting World Health Organization recommendations for fruit and vegetable intake. CONCLUSION: We found an unexpected positive association between DASH score for fruit intake and blood pressure parameters and hypertension among Vietnamese adults. More research is needed in this population to understand the relationship between vegetable and fruit intake with hypertension before a firm conclusion and recommendation are made.

Knowledge and Determinants of Health Consequences of Cigarette Smoking Among Vietnamese Adults, 2015.

Background. Vietnam was one of 16 countries that implemented the second round of the GATS (Global Adult Tobacco Survey) in 2015. Aims. To assess knowledge and determinants of health consequences of different types of cigarette smoking among Vietnamese adults. Methods. A cross-sectional study among adults aged 15 years combined with using 15% of the master sample from the national sampling frame of the population and housing census was conducted. Multilevel analysis using Poisson regression was undertaken. Results. Knowledge on the health consequences of cigarette smoking has not significantly improved in the GATS-2015. Adults believe that active smoking had more of an impact on health than secondhand smoking and 24.5% and 43% of them answered that smoking light and e-cigarettes, respectively, causes less harm than regular cigarettes, and 17.3% and 18.1% of adults are not aware of the difference between them, respectively. Conclusion. Household and community's role had little impact in the adult knowledge of smoking health consequences. It is necessary to disseminate information on the "other tobacco product" and to improve public knowledge on specific health consequences, to enhance household and community's role in conveying health education messages to individuals.

Multilocus sequence typing of Cryptococcus neoformans var. grubii from Laos in a regional and global context.

Cryptococcosis causes approximately 180 000 deaths each year in patients with human immunodeficiency virus (HIV). Patients with other forms of immunosuppression are also at risk, and disease is increasingly recognized in apparently immunocompetent individuals. Cryptococcus neoformans var. grubii, responsible for the majority of cases, is distributed globally. We used the consensus ISHAM Multilocus sequence typing (MLST) scheme to define the population structure of clinical C. neoformans var. grubii isolates from Laos (n = 81), which we placed into the global context using published MLST data from other countries (total N = 1047), including a reanalysis of 136 Vietnamese isolates previously reported. We observed a phylogeographical relationship in which the Laotian population was similar to its neighbor Thailand, being dominated (83%) by Sequence Types (ST) 4 and 6. This phylogeographical structure changed moving eastwards, with Vietnam's population consisting of an admixture of isolates dominated by the ST4/ST6 (35%) and ST5 (48%) lineages. The ST5 lineage is the predominant ST reported from China and East Asia, where it accounts for >90% of isolates. Analysis of genetic distance (Fst) between different populations of C. neoformans var. grubii supports this intermediate structure of the Vietnamese population. The pathogen and host diversity reported from Vietnam provide the strongest epidemiological evidence of the association between ST5 and HIV-uninfected patients. Regional anthropological genetic distances suggest diversity in the C. neoformans var. grubii population across Southeast Asia is driven by ecological rather than human host factors. Where the ST5 lineage is present, disease in HIV-uninfected patients is to be expected.

Spontaneous Otoacoustic Emissions Reveal an Efficient Auditory Efferent Network.

Purpose: Understanding speech often involves processing input from multiple modalities. The availability of visual information may make auditory input less critical for comprehension. This study examines whether the auditory system is sensitive to the presence of complementary sources of input when exerting top-down control over the amplification of speech stimuli. Method: Auditory gain in the cochlea was assessed by monitoring spontaneous otoacoustic emissions (SOAEs), which are by-products of the amplification process. SOAEs were recorded while 32 participants (23 women, nine men; Mage = 21.13) identified speech sounds such as "ba" and "ga." The speech sounds were presented either alone or with complementary visual input, as well as in quiet or with 6-talker babble. Results: Analyses revealed that there was a greater reduction in the amplification of noisy auditory stimuli compared with quiet. This reduced amplification may aid in the perception of speech by improving the signal-to-noise ratio. Critically, there was a greater reduction in amplification when speech sounds were presented bimodally with visual information relative to when they were presented unimodally. This effect was evidenced by greater changes in SOAE levels from baseline to stimuli presentation in audiovisual trials relative to audio-only trials. Conclusions: The results suggest that even the earliest stages of speech comprehension are modulated by top-down influences, resulting in changes to SOAEs depending on the presence of bimodal or unimodal input. Neural processes responsible for changes in cochlear function are sensitive to redundancy across auditory and visual input channels and coordinate activity to maximize efficiency in the auditory periphery.

Top-Down Cognitive and Linguistic Influences on the Suppression of Spontaneous Otoacoustic Emissions.

Auditory sensation is often thought of as a bottom-up process, yet the brain exerts top-down control to affect how and what we hear. We report the discovery that the magnitude of top-down influence varies across individuals as a result of differences in linguistic background and executive function. Participants were 32 normal-hearing individuals (23 female) varying in language background (11 English monolinguals, 10 Korean-English late bilinguals, and 11 Korean-English early bilinguals), as well as cognitive abilities (working memory, cognitive control). To assess efferent control over inner ear function, participants were presented with speech-sounds (e.g., /ba/, /pa/) in one ear while spontaneous otoacoustic emissions (SOAEs) were measured in the contralateral ear. SOAEs are associated with the amplification of sound in the cochlea, and can be used as an index of top-down efferent activity. Individuals with bilingual experience and those with better cognitive control experienced larger reductions in the amplitude of SOAEs in response to speech stimuli, likely as a result of greater efferent suppression of amplification in the cochlea. This suppression may aid in the critical task of speech perception by minimizing the disruptive effects of noise. In contrast, individuals with better working memory exert less control over the cochlea, possibly due to a greater capacity to process complex stimuli at later stages. These findings demonstrate that even peripheral mechanics of auditory perception are shaped by top-down cognitive and linguistic influences.

Language Experience Changes Audiovisual Perception.

Can experience change perception? Here, we examine whether language experience shapes the way individuals process auditory and visual information. We used the McGurk effect—the discovery that when people hear a speech sound (e.g., “ba”) and see a conflicting lip movement (e.g., “ga”), they recognize it as a completely new sound (e.g., “da”). This finding suggests that the brain fuses input across auditory and visual modalities demonstrates that what we hear is profoundly influenced by what we see. We find that cross-modal integration is affected by language background, with bilinguals experiencing the McGurk effect more than monolinguals. This increased reliance on the visual channel is not due to decreased language proficiency, as the effect was observed even among highly proficient bilinguals. Instead, we propose that the challenges of learning and monitoring multiple languages have lasting consequences for how individuals process auditory and visual information.

Linguistic Predictors of Cultural Identification in Bilinguals.

Most of the world's population has knowledge of at least two languages. Many of these bilinguals are also exposed to and identify with at least two cultures. Because language knowledge enables participation in cultural practices and expression of cultural beliefs, bilingual experience and cultural identity are interconnected. However, the specific links between bilingualism and cultural identity remain largely unidentified. The current study examined which aspects of bilingualism relate to identification with first- and second-language cultures. Two-hundred-and-nine bilinguals completed a questionnaire probing linguistic background and cultural affiliations. Regression analyses indicated that cultural identification was predicted by age of language acquisition, language proficiency, foreign-accentedness, and contexts of long-term language immersion and current language exposure. Follow-up analyses revealed that the language-culture relations were mediated by the age and manner in which the second language was acquired. These findings are situated within a proposed framework of bilingual cultural identity. By identifying features of bilingualism that are relevant for cultural identity, the current research increases our understanding of the relationship between language and culture.

Catecholamine-Dependent ß-Adrenergic Signaling in a Pluripotent Stem Cell Model of Takotsubo Cardiomyopathy.

BACKGROUND: Takotsubo syndrome (TTS) is characterized by an acute left ventricular dysfunction and is associated with life-threating complications in the acute phase. The underlying disease mechanism in TTS is still unknown. A genetic basis has been suggested to be involved in the pathogenesis. OBJECTIVES: The aims of the study were to establish an in vitro induced pluripotent stem cell (iPSC) model of TTS, to test the hypothesis of altered ß-adrenergic signaling in TTS iPSC-cardiomyocytes (CMs), and to explore whether genetic susceptibility underlies the pathophysiology of TTS. METHODS: Somatic cells of patients with TTS and control subjects were reprogrammed to iPSCs and differentiated into CMs. Three-month-old CMs were subjected to catecholamine stimulation to simulate neurohumoral overstimulation. We investigated ß-adrenergic signaling and TTS cardiomyocyte function. RESULTS: Enhanced ß-adrenergic signaling in TTS-iPSC-CMs under catecholamine-induced stress increased expression of the cardiac stress marker NR4A1; cyclic adenosine monophosphate levels; and cyclic adenosine monophosphate-dependent protein kinase A-mediated hyperphosphorylation of RYR2-S2808, PLN-S16, TNI-S23/24, and Cav1.2-S1928, and leads to a reduced calcium time to transient 50% decay. These cellular catecholamine-dependent responses were mainly mediated by ß1-adrenoceptor signaling in TTS. Engineered heart muscles from TTS-iPSC-CMs showed an impaired force of contraction and a higher sensitivity to isoprenaline-stimulated inotropy compared with control subjects. In addition, altered electrical activity and increased lipid accumulation were detected in catecholamine-treated TTS-iPSC-CMs, and were confirmed by differentially expressed lipid transporters CD36 and CPT1C. Furthermore, we uncovered genetic variants in different key regulators of cardiac function. CONCLUSIONS: Enhanced ß-adrenergic signaling and higher sensitivity to catecholamine-induced toxicity were identified as mechanisms associated with the TTS phenotype. (International Takotsubo Registry [InterTAK Registry] [InterTAK]; NCT01947621).

Comparative genomics of Cryptococcus neoformans var. grubii associated with meningitis in HIV infected and uninfected patients in Vietnam.

The vast burden of cryptococcal meningitis occurs in immunosuppressed patients, driven by HIV, and is caused by Cryptococcus neoformans var. grubii. We previously reported cryptococcal meningitis in Vietnam arising atypically in HIV uninfected, apparently immunocompetent patients, caused by a single amplified fragment length polymorphism (AFLP) cluster of C. neoformans var. grubii (VNIγ). This variant was less common in HIV infected individuals; it remains unclear why this lineage is associated with apparently immunocompetent patients. To study this host tropism we aimed to further our understanding of clinical phenotype and genomic variation within Vietnamese C. neoformans var. grubii. After performing MLST on C. neoformans clinical isolates we identified 14 sequence types (STs); ST5 correlated with the VNIγ cluster. We next compared clinical phenotype by lineage and found HIV infected patients with cryptococcal meningitis caused by ST5 organisms were significantly more likely to have lymphadenopathy (11% vs. 4%, p = 0.05 Fisher's exact test) and higher blood lymphocyte count (median 0.76 versus 0.55 X109 cells/L, p = 0.001, Kruskal-Wallis test). Furthermore, survivors of ST5 infections had evidence of worse disability outcomes at 70 days (72.7% (40/55) in ST5 infections versus 57.1% (52/91) non-ST5 infections (OR 2.11, 95%CI 1.01 to 4.41), p = 0.046). To further investigate the relationship between strain and disease phenotype we performed genome sequencing on eight Vietnamese C. neoformans var. grubii. Eight genome assemblies exhibited >99% nucleotide sequence identity and we identified 165 kbp of lineage specific to Vietnamese isolates. ST5 genomes harbored several strain specific regions, incorporating 19 annotated coding sequences and eight hypothetical proteins. These regions included a phenolic acid decarboxylase, a DEAD-box ATP-dependent RNA helicase 26, oxoprolinases, a taurine catabolism dioxygenase, a zinc finger protein, membrane transport proteins and various drug transporters. Our work outlines the complexity of genomic pathogenicity in cryptococcal infections and identifies a number of gene candidates that may aid the disaggregation of the pathways associated with the pathogenesis of Cryptococcus neoformans var. grubii.

Repetition reduction during word and concept overlap in bilinguals.

In natural conversation, speakers often mention the same referents multiple times. While referents that have been previously mentioned are produced with less prominence than those that have not been mentioned, it is unclear whether prominence reduction is due to repetition of concepts, words, or a combination of the two. In the current study, we dissociate these sources of repetition by examining bilingual speakers, who have more than one word for the same concept across their two languages. Three groups of Korean-English bilinguals (balanced, English-dominant, and Korean-dominant) performed an event description task involving repetition of referents within a single language (i.e., repetition of word and concept) or across languages (i.e., repetition of concept only). While balanced bilinguals reduced prominence both within and across languages, unbalanced bilinguals only reduced prominence when repetition occurred within a language. These patterns suggest that the degree to which words and concepts are linked within a speaker's language system determines the source of repetition reduction.

Repetition reduction: lexical repetition in the absence of referent repetition.

Compared to words that are new to a discourse, repeated words are produced with reduced acoustic prominence. Although these effects are often attributed to priming in the production system, the locus of the effect within the production system remains unresolved because, in natural speech, repetition often involves repetition of referents and lexical items simultaneously. Therefore, repetition reduction could be due to repeated mention of a referent or to repetition of a word or referring expression. In our study, we use an event description task to test whether repetition reduction is due to repetition of lexical items or to repeated mention of referents. The results show that repeated lexical items lead to reduced duration and intensity even in the absence of referent repetition, whereas repeated referents lead to reduced intensity alone. The general pattern suggests that repetition reduction is due most strongly to repetition of the lexical item, rather than repeated mention of the referent.

Repetition is easy: why repeated referents have reduced prominence.

The repetition and the predictability of a word in a conversation are two factors that are believed to affect whether it is emphasized: predictable, repeated words are less acoustically prominent than unpredictable, new words. However, because predictability and repetition are correlated, it is unclear whether speakers lengthen unpredictable words to facilitate comprehension or whether this lengthening is the result of difficulties in accessing a new (nonrepeated) lexical item. In this study, we investigated the relationship between acoustic prominence, repetition, and predictability in a description task. In Experiment 1, we found that repeated referents are produced with reduced prominence, even when these referents are unexpected. In Experiment 2, we found that predictability and repetition both have independent effects on duration and intensity. However, word duration was primarily determined by repetition, and intensity was primarily determined by predictability. The results are most consistent with an account in which multiple cognitive factors influence the acoustic prominence of a word.

The xenoantibody response and immunoglobulin gene expression profile of cynomolgus monkeys transplanted with hDAF-transgenic porcine hearts.

BACKGROUND: Recent work has indicated a role for anti-Gal alpha 1-3Gal (Gal) and anti-non-Gal xenoantibodies in the primate humoral rejection response against human-decay accelerating factor (hDAF) transgenic pig organs. Our laboratory has shown that anti-porcine xenograft antibodies in humans and non-human primates are encoded by a small number of germline IgV(H) progenitors. In this study, we extended our analysis to identify the IgV(H) genes encoding xenoantibodies in immunosuppressed cynomolgus monkeys (Macaca fascicularis) transplanted with hDAF-transgenic pig organs. METHODS: Three immunosuppressed monkeys underwent heterotopic heart transplantation with hDAF porcine heart xenografts. Two of three animals were given GAS914, a poly-L-lysine derivative shown to bind to anti-Gal xenoantibodies and neutralize them. One animal rejected its heart at post-operative day (POD) 39; a second animal rejected the transplanted heart at POD 78. The third monkey was euthanized on POD 36 but the heart was not rejected. Peripheral blood leukocytes (PBL) and serum were obtained from each animal before and at multiple time points after transplantation. We analyzed the immune response by enzyme-linked immunosorbent assay (ELISA) to confirm whether anti-Gal or anti-non-Gal xenoantibodies were induced after graft placement. Immunoglobulin heavy-chain gene (V(H)) cDNA libraries were then produced and screened. We generated soluble single-chain antibodies (scFv) to establish the binding specificity of the cloned immunoglobulin genes. RESULTS: Despite immunosuppression, which included the use of the polymer GAS914, the two animals that rejected their hearts showed elevated levels of cytotoxic anti-pig red blood cell (RBC) antibodies and anti-pig aortic endothelial cell (PAEC) antibodies. The monkey that did not reject its graft showed a decline in serum anti-RBC, anti-PAEC, and anti-Gal xenoantibodies when compared with pre-transplant levels. A V(H)3 family gene with a high level of sequence similarity to an allele of V(H)3-11, designated V(H)3-11(cyno), was expressed at elevated levels in the monkey that was not given GAS914 and whose graft was not rejected until POD 78. IgM but not IgG xenoantibodies directed at N-acetyl lactosamine (a precursor of the Gal epitope) were also induced in this animal. We produced soluble scFv from this new gene to determine whether this antibody could bind to the Gal carbohydrate, and demonstrated that this protein was capable of blocking the binding of human serum xenoantibody to Gal oligosaccharide, as had previously been shown with human V(H)3-11 scFv. CONCLUSIONS: DAF-transgenic organs transplanted into cynomolgus monkeys induce anti-Gal and anti-non-Gal xenoantibody responses mediated by both IgM and IgG xenoantibodies. Anti-non-Gal xenoantibodies are induced at high levels in animals treated with GAS914. Antibodies that bind to the Gal carbohydrate and to N-acetyl lactosamine are induced in the absence of GAS914 treatment. The animal whose heart remained beating for 78 days demonstrated increased usage of an antibody encoded by a germline progenitor that is structurally related, but distinct from IGHV311. This antibody binds to the Gal carbohydrate but does not induce the rapid rejection of the xenograft when expressed at high levels as early as day 8 post-transplantation.

Improved assessment of graft function by echocardiography in cynomolgus monkey recipients of hDAF-transgenic pig cardiac xenografts.

BACKGROUND: The current practice of evaluating heterotopic heart xenografts by palpation allows only detection of severe graft dysfunction, which indicates terminal graft failure. Therefore, we evaluated whether echocardiography is a better method of detecting early graft dysfunction as a marker of rejection in abdominal pig heart xenografts in cynomolgus monkeys. METHODS: Six cynomolgus monkeys received heterotopic heart transplants from pig donors transgenic for human decay-accelerating factor (hDAF). Induction therapy consisted of either cyclophosphamide or rabbit anti-thymocyte globulin. Maintenance therapy consisted of cyclosporine or tacrolimus, steroids, and sodium mycophenolate or mycophenolate mofetil, GAS914 (alphaGal oligosaccharide containing glycoconjugate), and for some animals TP10 (soluble complement receptor type 1). Echocardiography was performed immediately after transplantation and 3 times a week after surgery. We scored contractility and measured left ventricular wall thickness. Impaired contractility or increased wall thickness were considered graft dysfunction and were treated with pulse steroids. Palpation score was recorded daily. We also obtained myocardial biopsy specimens. RESULTS: Palpation score remained at 4 out of 4 in all animals until 2 to 5 days before final graft failure, whereas echocardiography detected several episodes of impaired graft function, either decreased left ventricular contractility or increased left ventricular wall thickness before graft failure. Treatment with pulse steroids improved graft function only during early episodes of graft impairment. Final graft failure was steroid resistant and caused by severe vascular rejection. CONCLUSIONS: Echocardiography is a better method of assessing graft dysfunction than is palpation. Therefore, echocardiography may detect early rejection episodes of heterotopic heart xenografts in non-human primates.